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Why image RNA?

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Numerous non-coding RNAs such lncRNAs, snRNAs, circRNAs or miRNAs are constantly identified hence their functional characterization is of great importance.

Learn how current aptamer-based RNA imaging and analysis strategies contribute to characterization of a specific RNA.

RNA aptamers are specific RNA sequences that can be detected by small-molecule dyes. Dye fluorescence is significantly enhanced upon complex formation thus aptamers are versatile tags for imaging and functional analysis of coding and non-coding RNA both in vitro and in live cells[1-4] (Tab. 1). Attachement of a specific aptamer sequence to a RNA of interest can be performed e.g. by in vitro transcription.

Table 1: Application of light-up aptamers in imaging & analysis of coding and non-coding RNA.

RNA Target Application Aptamer/Dye Combination Ref.
mRNA Live cell imaging (E. coli) of specific endogenous mRNA Spinach derivative/DFHBI [5]
Live cell imaging (mammalian cells) of specific endogenous mRNA Broccoli & Spinach derivatives/DFHBI-1T [6]
lncRNA Live cell imaging (mammalian cells) of aptamer-tagged lncRNA Broccoli derivative/DFHBI-1T [7]
miRNA miRNA detection in vitro Spinach derivative/DFHBI [8]
Spinach derivatives/DFHBI-1T [9]
snoRNA Live cell imaging (yeast) of aptamer-tagged snoRNA Broccoli derivative/DFHBI-1T [10]
tRNA Live cell imaging (E. coli) of ribosome activity Spinach derivative/DFHBI [11]
snoRNA/tRNA/5S RNA Live cell imaging (mammalian cells) and quantification of transcription Corn/DFHO [12]
 

Selected References:

[1] Neubacher et al. (2019) RNA Structure and Cellular Applications of Fluorescent Light-Up Aptamers. Angew. Chem. Int. Ed. 58:1266.
[2] Ouellet (2016) RNA Fluorescence with Ligth-Up Aptamers. Front. Chem 4:29.
[3] George et al. (2018) Intracelluar RNA-tracking methods. Open Biol 8:180104.
[4] Dologosheina et al. (2016) Fluorophore Binding RNA aptamers and their application. Wires RNA 7 (6):843.
[5] Ong et al. (2017) Live cell imaging of endogenous mRNA using RNA-based Fluorescence „Turn-On“ Probe. ACS Chem Biol 12 (1):200.
[6] Wang et al. (2019) Visualizing mRNA in live mammalian cells. Methods 2023 (18):30287.
[7] O‘Leary et al. (2017) Long non-coding RNA PARTICLEbridges histone and DNA methylation SCIENTIFIC Reports 7:1790.
[8] Zhou et al. (2019) Cascade transcription amplification of RNA aptamer for ultrasensitive microRNA detection. Anal. Chem. 91:5295.
[9] Ying et al. (2018) Spinach-based fluorescent light-up biosensors for multiplexed and label-free detection of microRNAs. Chem. Commun. 54:3010.
[10] Zinskieet al. (2018) Live-cell imaging of small nucleolar RNA tagged with the broccoli aptamer in yeast. Chem. Commun. 54:3010.
[11] Masuda et al. (2017) A genetically encoded fluorescent tRNA is active in live cell protein synthesis. Nucleic Acids Res. 45 (7):4081.
[12] Warner et al. (2017) A homodimer interface without base pairs in an RNA mimic of red fluorescent protein. Nat. Chem. Biol. 13 (11):1195.